Measurements Of Fluorescence Spectra Research Articles

A sensitive and reliable method for the quantitative determination of hydrogen peroxide produced by microalgae cells.

One of the reactive forms of oxygen is hydrogen peroxide (H2O2), which has been investigated as a key component of growth processes and stress responses. Different methods for the determination of H2O2 production by animal and bacterial cells exist; however, its detection in algal cell cultures is more complicated due to the presence of photosynthetic pigments in the cells and the complex structure of cell walls. Considering these issues, a reliable, quick, and simple method for H2O2 detection is needed in phycological research. The aim of this methodological study was to optimize an Amplex UltraRed method for the fluorometric detection of H2O2 produced by microalgae cells, using a wild-type strain of Chlamydomonas reinhardtii as a model. The results showed that (i) potassium phosphate is the most suitable reaction buffer for this method, (ii) a 560 nm wavelength variant is the most appropriate as the excitation wavelength for fluorescence spectra measurement, (iii) a 50:50 ratio for the reaction mixture to sample was the most suitable, (iv) the fluorescence signal was significantly influenced by the density of the microalgae biomass, and (v) sample fortification with H2O2 allowed for an increase of the method's reliability and repeatability. The proposed protocol of the Amplex UltraRed method for the fluorometric detection of H2O2 produced by microalgae cells can yield a sensitive and accurate determination of the content of the test compound, minimizing measurement errors, eliminating chlorophyll autofluorescence problem, and compensating for the matrix effect. This method can be applied to the study of other microalgae species.

An in situ approach for validation of canopy chlorophyll fluorescence radiative transfer models using the full emission spectrum

The intensity and spectral properties of solar-induced chlorophyll fluorescence (SIF) carry valuable information on plant photosynthesis and productivity, but are also influenced by leaf and canopy structure. Physically based models provide a quantitative means to investigate how SIF intensity and spectra propagate and scale from the photosystem to the leaf and to the canopy levels. However, the validation of canopy SIF models is limited by the lack of methods that combine direct, independent, and complementary measurements of the full fluorescence spectrum at the leaf and canopy levels. Here, we propose a novel validation approach that combines in situ measurements of leaf and canopy fluorescence spectra. The approach is demonstrated with measurements in a rice crop at two contrasting stages of canopy development. We measured leaf reflectance, transmittance, and fluorescence spectra in situ, and subsequently inverted leaf structural and biochemical parameters and determined the leaf fluorescence quantum efficiency (FQE) using the Fluspect-Cx model. Two FQE inversion methods (Inversion-IIA and Inversion-IIB) were tested for the forward simulation of leaf fluorescence spectra. Leaf fluorescence spectra were then scaled up to the canopy level using 1D, 2D, and 3D radiative transfer schemes (SCOPE, mSCOPE, and DART), and compared with the direct canopy fluorescence spectral observations measured under red, green, blue, and white illumination. The validation results demonstrate that accounting for 3D canopy structure, as in the DART model, is critical to successfully scale the fluorescence spectrum from the leaf to the canopy level, whereas 1D SCOPE or even 2D mSCOPE were unable to fully reproduce the canopy fluorescence spectra. The results also demonstrate that the Inversion-IIB method matches relatively well the measurements with mean relative absolute errors (MRAE) of 20 %, 37 %, and 43 % versus Inversion-IIA with mean relative absolute errors (MRAE) of 62 %, 100 %, and 108 % for DART, mSCOPE, and SCOPE, respectively. We suggest that our validation approach is transferable to other plant species and canopy geometries, providing a means to standardize and evaluate the performance of canopy SIF models and improve our understanding of canopy SIF observations.

Observation of Space-Dependent Rotational Doppler Shifts with a Single Ion Probe.

We present an experiment investigating the rotational Doppler effect using a single trapped ion excited by two copropagating vortex laser beams. The setup isolates the azimuthal gradients of the fields, eliminating longitudinal and curvature effects. We provide a detailed characterization of the phenomenon by deterministically positioning a single ion across the beams and measuring fluorescence spectra with sharp "dark resonances" whose features depend on the angular velocity of the ion and the difference of optical orbital angular momentum between the two beams. The interpretation of the measurements is supported by numerical simulations and by a simplified analytical model. Our results reveal key properties of the rotational Doppler effect, showing that it increases approaching the center of the beam and that it is independent of the waist of the beam. This offers insights into the feasibility of superkicks or super-Doppler shifts for sensing and manipulating atomic motion transverse to the beams' propagation direction.

Effect of InP/ZnS quantum dots aggregation on the kinetics of birefringence recorded in thin azopolymer composite films

The article describes the preparation of new thin film composite materials based on the azopolymer (poly[1-[4-(3-carboxy-4-hydroxyphenylazo)benzenesulfonamido]-1,2-ethanediyl, sodium salt]) doped with three different concentration of InP/ZnS quantum dots with size of 7.5 nm. Clusters of aggregated InP/ZnS were microscopically observed in the fabricated composite samples. Birefringence (Δn) was induced in the films at two different wavelengths of the pump laser (355 nm and 444 nm) and the obtained higher values of Δnmax for the samples doped with quantum dots up to 2 wt% in comparison to the non-doped film and the sample with the highest nanocrystals concentration are discussed, based on the measured fluorescence spectra, in terms of possible local energy transfer between the azobenzene chromophores and the InP/ZnS quantum dots.

Dose Optimization of Intravenous Indocyanine Green for Malignant Lung Tumor Localization.

Background: Intravenously administered indocyanine green (ICG) accumulates in lung tumors, facilitating their detection via a fluorescence spectrum measurement. This method aids in identifying tumor locations that are invisible to the naked eye. We aim to determine the optimal ICG dose and administration method for accurate tumor identification during lung resection surgeries, utilizing a novel ICG fluorescence spectroscopy system for precise tumor localization. Materials and Methods: ICG should be dissolved in the provided solution or distilled water and administered intravenously approximately 24 h before surgery, beginning with an initial dose of 0.5 mg/kg. If the tumor detection rate is insufficient, the dose may be gradually increased to a maximum of 5.0 mg/kg to determine the optimal dosage for effective tumor detection. This fluorescence spectroscopy during surgery may reveal additional lesions that remain undetected in preoperative assessments. The primary endpoint includes the correct diagnostic rate of tumor localization. The secondary endpoints include the measurement of the intraoperative ICG fluorescence spectral intensity in lung tumors, the assessment of the operability and safety of intraperitoneal ICG administrations, the measurement of the ICG fluorescence spectral intensity in surgical specimens, the comparison of the spectral intensity in lung tissues during collapse and expansion, the correlation between ICG camera images and fluorescence spectral intensity, and the comparison of fluorescence analysis results with histopathological findings. The trial has been registered in the jRCT Clinical Trials Registry under the code jRCTs011230037. Results and Conclusions: This trial aims to establish an effective methodology for localizing and diagnosing malignant lung tumors, thereby potentially improving surgical outcomes and refining treatment protocols.

Оn the aggregation of polycationic photosensitizer upon binding to Gram-negative bacteria

Polycationic photosensitizers (PS) are not susceptible to aggregation in solutions, but their high local concentrations in Gram-negative bacteria can be sufficient for aggregation and reduced effectiveness of antibacterial photodynamic treatment. By measuring fluorescence spectra and kinetics we were able to evaluate the degree of aggregation of polycationic PS ZnPcChol8 in Gram-negative bacteria E. coli K12 TG1. Binding of ZnPcChol8 to E. coli K12 TG1 leads to an appearance of groups of molecules with shorter PS fluorescence lifetime, a decrease in fluorescence intensity and a shift in the fluorescence spectral maximum. However, we evaluated that about 88% of the fluorescing PS molecules in the bacteria were in an unaggregated state, which indicates only a small reduction in the generation of reactive oxygen species.

Synthesis of bis-(benzhydryloxy) substituted axially silicon(IV) phthalocyanine: investigation of photophysical, photochemical, and computational electronic properties

Synthesis of axially disubstituted silicon(IV) phthalocyanine bearing benzhydryloxy groups obtained by reaction of SiPcCl2 with diphenylmethanol is reported. Characterization of the compound was performed by 1H and 13C NMR, UV–Vis, mass spectrometry, and FT-IR spectroscopy. Photophysical and photochemical properties were investigated. Fluorescence spectra measurements were performed in DMSO as solvent. A preferred chemical method was used to determine the amount of singlet oxygen production. Optimizations were calculated for the single-molecule Si-Pc with the 6–311 G (d, p) basis set at the B3LYP and B3PW91 levels of density functional theory (DFT). The charge transfer (or) charge delocalization between intramolecular donor–acceptor groups was determined by NBO analysis. Furthermore, molecular electrostatic potential, Fukui functions, electrophilic and nucleophilic character maps were calculated. The electronic structure and energy parameters of the investigated compound were determined to be ΔE calc = 2.19 eV by DFT calculation and ΔE opt = 1.84 by experimental UV spectrum.

Real-time pollen identification using holographic imaging and fluorescence measurements

Abstract. Over the past few years, a diverse range of automatic real-time instruments has been developed to respond to the needs of end users in terms of information about atmospheric bioaerosols. One of them, the SwisensPoleno Jupiter, is an airflow cytometer used for operational automatic bioaerosol monitoring. The instrument records holographic images and fluorescence information for single aerosol particles, which can be used for identification of several aerosol types, in particular different pollen taxa. To improve the pollen identification algorithm applied to the SwisensPoleno Jupiter and currently based only on the holography data, we explore the impact of merging fluorescence spectra measurements with holographic images. We demonstrate, using measurements of aerosolised pollen, that combining information from these two sources results in a considerable improvement in the classification performance compared to using only a single source (balanced accuracy of 0.992 vs. 0.968 and 0.878). This increase in performance can be ascribed to the fact that often classes which are difficult to resolve using holography alone can be well identified using fluorescence and vice versa. We also present a detailed statistical analysis of the features of the pollen grains that are measured and provide a robust, physically based insight into the algorithm's identification process. The results are expected to have a direct impact on operational pollen identification models, particularly improving the recognition of taxa responsible for respiratory allergies.

Study the effect of nano NiCo2O4 doping on the optical and dielectric properties of PMMA/PEO/MWCNTs

In this study, NiCo2O4 nanoparticles were prepared using a hydrothermal process and examined by x-ray diffraction. After that, using the casting technique, blends of polymethyl methacrylate (PMMA)/poly(ethylene oxide) (PEO) were loaded with multi-walled carbon nanotubes (MWCNTs) and different amounts (x) of NiCo2O4 nanoparticles. The obtained blends were characterized using X-ray diffraction and scanning electron microscopy techniques. Utilizing diffuse reflectance measurements, various optical parameters, including energy gap, refractive index, extinction coefficient, optical dielectric constant, optical conductivity, and nonlinear optical parameters, were investigated. Following incorporation of MWCNTs/NiCo2O4 in the blend, the transmittance of the PMMA/PEO blend (90 %) displayed a drop reaching 21 %, the reflectivity and refractive index increased. The direct/indirect optical band gap values of PMMA/PEO (5.07/4.8 eV) decreased attaining 4.96/4.54 eV for the blend with 2.5 wt% NiCo2O4. The optical conductivity greatly enhanced, and the linear and nonlinear optical parameters increased as the amount of NiCo2O4 doping increased. The dispersion parameters for all blends were also calculated. Fluorescence spectra (FL) measurements revealed a reduction in the FL intensity with increasing the NiCo2O4 content (x). Through the dielectric measurements, the impact of increasing NiCo2O4 content on the ac conductivity, dielectric properties, energy density, and electric modulus was analyzed for all blends. Remarkably, a significant increase in AC conductivity was obtained upon loading with 1.5 wt% NiCo2O4. Overall, variation in the dielectric and optical features of the PMMA/PEO/MWCNTs/x wt.% NiCo2O4 blends makes them appropriate for optoelectronic and energy storage applications.

Diphenolic boldine, an aporphine alkaloid: inhibitory effect evaluation on α-glucosidase by molecular dynamics integrating enzyme kinetics

Screening α-glucosidase inhibitors with novel structures is an important field in the development of anti-diabetic drugs due to their application in postprandial hyperglycemia control. Boldine is one of the potent natural antioxidants with a wide range of pharmacological activities. Virtual screening and biochemical inhibition kinetics combined with molecular dynamics simulations were conducted to verify the inactivation function of boldine on α-glucosidase. A series of inhibition kinetics and spectrometry detections were conducted to analyze the α-glucosidase inhibition. Computational simulations of molecular dynamics/docking analyses were conducted to detect boldine docking sites’ details and evaluate the key binding residues. Boldine displayed a typical reversible and mixed-type inhibition manner. Measurements of circular dichroism and fluorescence spectrum showed boldine changed the secondary structure and loosened the tertiary conformation of target α-glucosidase. The computational molecular dynamics showed that boldine could block the active pocket site through close interaction with binding key residues, and two phenolic hydroxyl groups of boldine play a core function in α-glucosidase inhibition via ligand binding. This investigation reveals the boldine function on interaction with the α-glucosidase active site, which provides a new inhibitor candidate. Communicated by Ramaswamy H. Sarma

Excited-state proton transfer based fluorescence in Kaempferol powder and solutions with different concentrations

Kaempferol (KMP) is one of the most common flavonoids, currently being extensively studied for its numerous beneficial health effects. Here we study the fluorescence (FL) emission of KMP powder and of its solutions prepared using different types of solvents (polar and non-polar). In the spectra of KMP powder and KMP solutions with high concentration, the same FL peak with maximum at 1.9 eV is observed. Another FL peak, at higher energy of 2.45 eV, emerges in solutions, its relative intensity increases with decreasing solution concentration. The FL emission of solutions with lowest concentration displays only that peak. To calculate characteristic energies of absorption and emission of KMP molecule in vacuum and in solutions we use time-dependent density functional theory. Comparing the results of computations with measured FL spectra, we associate the FL band at 1.9 eV with the emission due to excited state intramolecular transfer of the proton of –OH5 hydroxyl group. The FL emission at 2.45 eV is related to the –OH3 proton transfer. We measure the FL spectra of KMP powder using two different excitation energies, 3.06 eV and 2.33 eV, and find that its FL spectrum depends on the excitation energy. To understand that dependence, we compare the FL spectra of KMP and Q monohydrate powders. We consider the excited state intermolecular transfer of the proton from –OH3′ hydroxyl group to a neighboring molecule in Q crystal and calculate the energy corresponding to the emission of the resulted anion of Q molecule. The spectral feature at 1.69 eV observed only in the FL spectrum of Q hydrate is attributed to the Q anion FL emission.

Dual-state emission of D-A-D type benzothiadiazole derivatives for the sensitive detection of amine compounds

Organic light-emitting materials with dual-state emission (DSE) in both solution and solid states have attracted widely attention because of their promising applications in diverse areas. In this work, five benzothiadiazole-based D-A-D type luminogens BT2Ph, BT2Na, BT2Pa, BT2An and BT2Py were concisely synthesized by introducing different π–conjugated rigid groups (benzene, naphthalene, phenanthrene, anthracene and pyrene ring). As a result of their twisted conformation and effective conjugation structure, these five dual-state emission molecules exhibit intense photoluminescence in solution and solid state. The reason for these luminogens with DSE behavior was further explained by the twisted intramolecular charge-transfer mechanism based on the variable temperature fluorescence spectrum measurement and theoretical calculation with time-dependent density functional theory (TD-DFT). Moreover, these DSE luminogens which possess excellent photostability and obvious solvatofluorochromic properties were multifunctional fluorescent probes for the visual detection of amine compounds (aniline, cyclohexylamine, and ammonia). A charge transfer mechanism between luminogens and amine compounds was found to be the cause of fluorescence quenching. Thus, these DSE molecules may have applications in anti-counterfeiting and fluorescent sensors.

Spectroscopic and molecular docking study of three kinds of cinnamic acid interaction with pepsin

In this work, under simulated physiological conditions (pH = 2.2, glycine hydrochloric acid buffer solution), the interactions of cinnamic acid (CA), m-hydroxycinnamic acid (m-CA) and p-hydroxycinnamic acid (p-CA) with pepsin were studied by fluorescence spectroscopy, ultraviolet–visible absorption spectroscopy, circular dichroism (CD) spectroscopy, Fourier transform infrared spectroscopy (FTIR), molecular docking and molecular dynamic simulation (MD). The spectrogram results showed that these three kinds of CA had a strong ability to quench the intrinsic fluorescence of pepsin, and the quenching effects were obvious with the increase of concentration of these three kinds of molecules. The quenching mechanism of CA, m-CA and p-CA on the fluorescence of pepsin was static quenching. In addition, a stable complex was formed between three kinds of CA with pepsin. Thermodynamic data and docking information suggested that three kinds of CA combine with pepsin were mainly driven by electrostatic force and hydrogen bond. The binding constant and the number of binding sites were determined. The interaction of CA, m-CA and p-CA with pepsin was spontaneous, and accompanied by non-radiative energy transfer. The results from CD, FTIR, UV–Vis and synchronous fluorescence spectra measurements manifested that the secondary structure of pepsin was changed by the binding of three kinds of CA. The β-sheet of pepsin increased after the interaction with three kinds of CA. The assay results of pepsin activity showed that three kinds of CA led to a decrease in pepsin activity within the investigated concentrations. Molecular docking investigation revealed the formation of polar hydrogen bonds as well as hydrophobic interactions between three kinds of CA with pepsin, and the ligand within the binding pocket of pepsin. MD results implied the formation of a stable complex between three kinds of CA and pepsin. The research suggested that cinnamic acid and its derivatives could be a potential effect on the structure and properties of digestive enzyme.

Theoretical and experimental investigation of novel quinazoline derivatives: synthesis, photophysical, reactive properties, molecular docking and selective HSA biointeraction

Two new quinazoline derivatives (2a and 2b) were successfully synthesized in this work using the condensation technique in excellent yields. Using spectroscopic techniques and elemental analyses, the compounds were completely characterized. Density functional theory (DFT) computations have been used to examine the title compound’s reactive characteristics. Chemical reactivity was predicted using local reactive descriptors and molecule electrostatic potential. Additionally, Time dependent DFT (TD-DFT) simulations were used to examine the impact of solvents on the photophysical characteristics. The affinity of compounds 2a and 2b for human serum albumin (HSA) was further explored using several electronic spectroscopies. Through static mechanisms, both compounds reduce the intrinsic fluorescence of HSA. It is determined that the HSA-2b complex’s binding constant is significantly greater than the HSA-2a complex. The fluorescence spectrum measurements proved that the HSA underwent structural changes after interaction with these compounds. It was demonstrated by site marker competitive displacement studies that compounds 2a and 2b preferred to bind to site I in HSA subdomain IIA. Additionally, synchronised fluorescence spectra were utilized to analyze how HSA’s conformation changed after interacting with various substances. The molecular docking investigations of these compounds with the three critical HSA binding sites, comprising subdomains IIA, IIIA, and IB, further confirmed the experimental findings. The significant contact between the investigated compounds and HSA was supported by the docking simulations. Communicated by Ramaswamy H. Sarma

Impact of site-selective spectroscopy on laser cooling parameter characterization.

From laser design to optical refrigeration, experimentally measured fluorescence spectra are often utilized to obtain input parameters for predictive models. However, in materials that exhibit site-selectivity, the fluorescence spectra depend on the excitation wavelength employed to take the measurement. This work explores different conclusions that predictive models reach after inputting such varied spectra. Here, temperature-dependent site-selective spectroscopy is carried out on an ultra-pure Yb, Al co-doped silica rod fabricated by the modified chemical vapor deposition technique. The results are discussed in the context of characterizing ytterbium doped silica for optical refrigeration. Measurements made between 80 K and 280 K at several different excitation wavelengths yield unique values and temperature dependencies of the mean fluorescence wavelength. For the excitation wavelengths studied here, the variation in emission lineshapes ultimately lead to calculated minimum achievable temperatures (MAT) ranging between 151 K and 169 K, with theoretical optimal pumping wavelengths between 1030 nm and 1037 nm. Direct evaluation of the temperature dependence of the fluorescence spectra band area associated with radiative transitions out of the thermally populated 2F5/2 sublevel may be a better approach to identifying the MAT of a glass where site-selective behavior precludes unique conclusions.

Distinguishing Healthy and Carcinoma Cell Cultures Using Fluorescence Spectra Decomposition with a Genetic-Algorithm-Based Code.

In this paper, we analysed the steady state fluorescence spectra of cell suspensions containing healthy and carcinoma fibroblast mouse cells, using a genetic-algorithm-spectra-decomposition software (GASpeD). In contrast to other deconvolution algorithms, such as polynomial or linear unmixing software, GASpeD takes into account light scatter. In cell suspensions, light scatter plays an important role as it depends on the number of cells, their size, shape, and coagulation. The measured fluorescence spectra were normalized, smoothed and deconvoluted into four peaks and background. The wavelengths of intensities' maxima of lipopigments (LR), FAD, and free/bound NAD(P)H (AF/AB) of the deconvoluted spectra matched published data. In deconvoluted spectra at pH = 7, the fluorescence intensities of the AF/AB ratio in healthy cells was always higher in comparison to carcinoma cells. In addition, the AF/AB ratio in healthy and carcinoma cells were influenced differently by changes in pH. In mixtures of healthy and carcinoma cells, AF/AB decreases when more than 13% of carcinoma cells are present. Expensive instrumentation is not required, and the software is user friendly. Due to these attributes, we hope that this study will be a first step in the development of new cancer biosensors and treatments with the use of optical fibers.

Cross-section spectra and transient characteristics of Er3+ emissions in Gd3(Al, Ga)5O12 garnet single crystal

We measured fluorescence spectra of visible and infrared electronic transitions at 530 (2H11/2 → 4I15/2), 560 (4S3/2 → 4I15/2), 670 (4F9/2 → 4I15/2), 980 (4I11/2 → 4I15/2), 1530 (4I13/2 → 4I15/2) and 2700 nm (4I11/2 → 4I13/2) of Er3+ in Gd3(Al0.4Ga0.6)5O12 garnet single-crystal excited at the wavelength 520 nm (for the 980 nm emission) or 980 nm (for other emissions). From measured fluorescence spectrum, Er3+ emission cross-section spectrum was calibrated for each transition. Absorption cross-section spectra were further derived on the basis of McCumber equation, and compared with those spectra obtained from measured absorption spectrum on the basis of the Beer-Lambert law. The comparison showed that the results given by two methods can be regarded as same within the error. The transient features of visible and infrared emissions were also studied under pulse excitation. The results showed that, while the three IR emissions at 980, 1530 and 2700 nm all follow a mono-exponential function, both green and red 980-nm-upconverted emissions at 560 and 670 nm exhibit non-mono-exponentially decaying feature, and each decaying trace can be fitted by a sum of two exponential functions. The non-mono-exponential feature is related to cross relaxation caused by non-radiative short-distance energy transfer between clustered Er3+ sites.

The choice of spectral ranges of registration for the fluorescent method for detecting leaks in oil pipelines at an excitation wavelength of 355 nm

Mathematical modeling was carried out based on the experimentally measured fluorescence spectra of oil, vegetation and water to select the most effective spectral registration ranges for the fluorescent method for detecting oil leaks at an excitation wavelength of 355 nm. The results of mathematical modeling show that the probabilities of correct detection and false alarms for the problem of detecting oil leaks significantly depend on the type of oil and, accordingly, on the spectral channels selected for monitoring. For reliable detection of oil spills against the background of vegetation or water bodies, two or three spectral channels must be used. The highest probabilities of correct detection (>0.999) and small probabilities of false alarms (<0.04) can be achieved for oils with intensity maxima of laser-induced fluorescent radiation at wavelengths of ~ 420 and 550 nm (when using two spectral channels) and for oils with fluorescence intensity maximum at a wavelength of ~510 nm (when using three spectral channels). For oils with a maximum fluorescence intensity at a wavelength of about 475 nm (when using three spectral channels), the results are worse, although they remain acceptable (at a measurement noise of 10%, the probability of correct detection and false alarms, respectively, 0.94 and 0.11).

CFSA-AGD: An accurate crosstalk fluorescence spectroscopic decomposition method for identifying and quantifying FDOMs in aquatic environments

Fluorescent substances exist in various aquatic environments and other environmental media. It is a critical task to identify the components accurately and quantify their contents precisely. Based on the Crosstalk Fluorescence Spectroscopy Analysis (CFSA) model, a fluorescence spectroscopic decomposition using the Alternating Gradient Descent (AGD) algorithm is developed. By reducing the residual error of the model through alternating iterations, the CFSA-AGD method achieves unsupervised model training and automatic spectroscopic decomposition without extra experimental operations such as dilution or absorbance measurement, exempting from tedious modeling process. The objectives of this work are to validate that the CFSA-AGD method can comprehensively address the decomposition of fluorescence spectral crosstalk. Furthermore, the novel method is applied to the spectroscopic decomposition of natural FDOMs in aquatic environments as a standard tool. The spectral data analyzing the performance of this method is verified and compared with the conventional methods through the experiment on standard samples. The results indicate that CFSA-AGD has higher spectroscopic decomposition accuracy and gives more abundant information on the characteristic spectra with less residual error than parallel factor analysis. This means that the fluorescence spectra of natural FDOMs can be decomposed into the characteristic fluorescence emission spectra of single components with higher accuracy and the characteristic fluorescence absorption spectra that cannot be obtained by the conventional methods. Meanwhile, it improves the analytical precision of the contents (from R2 ≥ 0.9778 to R2 ≥ 0.9920) and reduces the ultimate residual error by two orders of magnitude (from 1.42 × 10−1 to 4.68 × 10−3) when the method is used to estimate the measured fluorescence spectra.

Spectroscopic investigations on fungal aspartic protease as target of gallic acid

Proteases are a major virulence factor in pathogenic fungi and can serve as a potential therapeutic target. The interaction of gallic acid (GA) with Aspartic fungal protease (PepA) was investigated using biophysical and in silico approaches. UV–Vis and fluorescence spectroscopy showed complex formation and static quenching of PepA by GA with Ka of 7.4 × 105 M−1 and stoichiometric binding site (n) of 1.67. CD-spectroscopy showed marked changes in helical content and synchronous fluorescence spectra measurements indicated significant changes in the microenvironment around tryptophan residues in the GA-PepA complex. Outcomes of Isothermal Titration Calorimetry (ITC) measurement and molecular modelling studies validated the spectroscopic results. The binding of GA to Human Serum albumin (HSA) was moderate (Ka = 1.9 × 103 M−1) and did not cause structural disruption of HSA. To conclude, gallic acid is strongly bound to fungal protease leading to structural disruption and inhibition whereas HSA structure was largely conserved. Gallic acid thus appears to be a potential therapeutic agent against fungal proteases.